Partial Prion Cross-Seeding between Fungal and Mammalian Amyloid Signaling Motifs.

In filamentous fungi, NLR-based signalosomes activate downstream membrane-targeting cell death-inducing proteins by a mechanism of amyloid templating. In the species Podospora anserina, two such signalosomes, NWD2/HET-S and FNT1/HELLF, have been described. An analogous system involving a distinct amyloid signaling motif, termed PP, was also identified in the genome of the species Chaetomium globosum and studied using heterologous expression in Podospora anserina The PP motif bears resemblance to the RIP homotypic interaction motif (RHIM) and to RHIM-like motifs controlling necroptosis in mammals and innate immunity in flies. We identify here a third NLR signalosome in Podospora anserina comprising a PP motif and organized as a two-gene cluster encoding an NLR and an HELL domain cell death execution protein termed HELLP. We show that the PP motif region of HELLP forms a prion we term [π] and that [π] prions trigger the cell death-inducing activity of full-length HELLP. We detect no prion cross-seeding between HET-S, HELLF, and HELLP amyloid motifs. In addition, we find that, like PP motifs, RHIMs from human RIP1 and RIP3 kinases are able to form prions in Podospora and that [π] and [Rhim] prions partially cross-seed. Our study shows that Podospora anserina displays three independent cell death-inducing amyloid signalosomes. Based on the described functional similarity between RHIM and PP, it appears likely that these amyloid motifs constitute evolutionarily related cell death signaling modules.IMPORTANCE Amyloids are ß-sheet-rich protein polymers that can be pathological or display a variety of biological roles. In filamentous fungi, specific immune receptors activate programmed cell death execution proteins through a process of amyloid templating akin to prion propagation. Among these fungal amyloid signaling sequences, the PP motif stands out because it shows similarity to the RHIM, an amyloid sequence controlling necroptotic cell death in mammals. We characterized an amyloid signaling system comprising a PP motif in the model species Podospora anserina, thus bringing to three the number of independent amyloid signaling cell death pathways described in that species. We then showed that human RHIMs not only propagate as prions in P. anserina but also partially cross-seed with fungal PP prions. These results indicate that, in addition to showing sequence similarity, the PP and RHIM motifs are at least partially functionally related, supporting a model of long-term evolutionary conservation of amyloid signaling mechanisms from fungi to mammals.

Characterization of cultural traits and fungicidal activity of strains belonging to the fungal genus Chaetomium.

AIMS: Compare and characterize Chaetomium strains with special regard to their potentialities as biocontrol agents. METHODS AND RESULTS: Twelve strains of the fungal genus Chaetomium from diverse ecological niches were identified as belonging to six different species. Large differences were observed between the strains with regard to temperature requirements for mycelial growth and pigmentation of culture filtrates. Culture filtrates and ethyl acetate extracts were assayed for fungicidal effects against important phytopathogens both on agar media and in multiwell plates. The samples from Chaetomium globosum were particularly active against Botrytis cinerea, Pyrenophora graminea and Bipolaris sorokiniana, while those from C. cochliodes and C. aureum were inhibitory towards Phytophthora infestans, and P. infestans and Fusarium culmorum respectively. To narrow down the active principle, the most promising extracts were separated by preparative HPLC and the resulting fractions tested in bioassays. Chaetoglobosins were identified as active compounds produced by C. globosum. CONCLUSIONS: The bioassays revealed C. aureum and C. cochliodes as promising candidates for use in biocontrol. Both showed remarkably good activity against the prominent plant pathogen P. infestans. SIGNIFICANCE AND IMPACT OF THE STUDY: We provide the first systematic study comparing six different Chaetomium species with regard to their use as biocontrol agents.

Conventional detection and quantification real-time PCR of the pks-1 gene of Chaetomium globosum.

Chaetomium globosum is known as a potential biocontrol indicator against various soil and seedborne pathogens. Precise data are necessary for population monitoring of C. globosum for its effective use in agriculture. A sequence-characterized amplified region marker has been applied for the detection of this biocontrol agent, which will help to detect C. globosum at the site of its application. Out of 17 isolates of C. globosum, only 8 isolates of C. globosum amplified a monomorphic band of 1,900 bp. C. globosum is known for producing chaetoglobosin A. The pks-1 gene is unique in C. globosum in that it is involved in chaetoglobosin A production, melanin formation, and sporulation. Real-time PCR of pks-1 was used to compare the expressions of the pks-1 gene and chaetoglobosin A biosynthesis and sporulation. It was found that the sporulation of C. globosum was associated with the levels of pks-1 gene expression; Cg2 isolate showed high expression of the pks-1 gene, 41.21%, and also produced a great number of spores and perithecia. The association between the pks-1 gene expression and chaetoglobosin A production was estimated. The Pks-1 gene was expressed by all C. globosum isolates except one isolate, C1, which is another species of Chaetomium. In addition, all C. globosum isolates produced chaetoglobosin A with different concentrations and did not express the same levels of pks-1. This finding may be a result of the solvent type used in the extraction.

Transcriptome analysis reveals the defense mechanism of cotton against Verticillium dahliae in the presence of the biocontrol fungus Chaetomium globosum CEF-082.

BACKGROUND: Verticillium wilt of cotton is a serious soil-borne disease that causes a substantial reduction in cotton yields. A previous study showed that the endophytic fungus Chaetomium globosum CEF-082 could control Verticillium wilt of cotton, and induce a defense response in cotton plants. However, the comprehensive molecular mechanism governing this response is not yet clear. RESULTS: To study the signalling mechanism induced by CEF-082, the transcriptome of cotton seedlings pretreated with CEF-082 was sequenced. The results revealed 5638 DEGs at 24 h post inoculation with CEF-082, and 2921 and 2153 DEGs at 12 and 48 h post inoculation with Verticillium dahliae, respectively. At 24 h post inoculation with CEF-082, KEGG enrichment analysis indicated that the DEGs were enriched mainly in the plant-pathogen interaction, MAPK signalling pathway-plant, flavonoid biosynthesis, and phenylpropanoid biosynthesis pathways. There were 1209 DEGs specifically induced only in cotton plants inoculated with V. dahliae in the presence of the biocontrol fungus CEF-082, and not when cotton plants were only inoculated with V. dahliae. GO analysis revealed that these DEGs were enriched mainly in the following terms: ROS metabolic process, H2O2 metabolic process, defense response, superoxide dismutase activity, and antioxidant activity. Moreover, many genes, such as ERF, CNGC, FLS2, MYB, GST and CML, that regulate crucial points in defense-related pathways were identified and may contribute to V. dahliae resistance in cotton. These results provide a basis for understanding the molecular mechanism by which the biocontrol fungus CEF-082 increases the resistance of cotton to Verticillium wilt. CONCLUSIONS: The results of this study showed that CEF-082 could regulate multiple metabolic pathways in cotton. After treatment with V. dahliae, the defense response of cotton plants preinoculated with CEF-082 was strengthened.

Crystal structures of Rea1-MIDAS bound to its ribosome assembly factor ligands resembling integrin-ligand-type complexes.

The Rea1 AAA+-ATPase dislodges assembly factors from pre-60S ribosomes upon ATP hydrolysis, thereby driving ribosome biogenesis. Here, we present crystal structures of Rea1-MIDAS, the conserved domain at the tip of the flexible Rea1 tail, alone and in complex with its substrate ligands, the UBL domains of Rsa4 or Ytm1. These complexes have structural similarity to integrin α-subunit domains when bound to extracellular matrix ligands, which for integrin biology is a key determinant for force-bearing cell-cell adhesion. However, the presence of additional motifs equips Rea1-MIDAS for its tasks in ribosome maturation. One loop insert cofunctions as an NLS and to activate the mechanochemical Rea1 cycle, whereas an additional ß-hairpin provides an anchor to hold the ligand UBL domains in place. Our data show the versatility of the MIDAS fold for mechanical force transmission in processes as varied as integrin-mediated cell adhesion and mechanochemical removal of assembly factors from pre-ribosomes.

Chaetomium atrobrunneum causing human eumycetoma: The first report.

In this communication, a case of black grain eumycetoma produced by the fungus C. atrobrunneum is reported. The patient was initially misdiagnosed with M. mycetomatis eumycetoma based on the grains' morphological and cytological features. However, further aerobic culture of the black grains generated a melanised fungus identified as C. atrobrunneum by conventional morphological methods and by internal transcribed spacer 2 (ITS2) ribosomal RNA gene sequencing. This is the first-ever report of C. atrobrunneum as a eumycetoma-causative organism of black grain eumycetoma. It is essential that the causative organism is identified to the species level, as this is important for proper patient management and to predict treatment outcome and prognosis.

Biological treatment of lignocellulosic biomass by Chaetomium globosporum: Process derivation and improved biogas production.

Chaetomium globosporum was isolated from aeolian soil samples collected from semi-arid locations in the state of Rajasthan, India. The efficiency of fungal strain for biological treatment of biomass to improve biogas yield was screened by estimating laccase enzyme activity under submerged fermentation system. Further, lignocellulosic biomass(s) wheat and pearl millet straw were subjected to biological treatment and subsequent increase in release of reducing sugar as compared to untreated straw was determined. Optimization of biological treatment conditions was accomplished by employing response surface methodology with Box Behnken design of experiments. Impact of three parameters (temperature, residence time, moisture content) was analyzed using three severity levels on output response of biological treatment in terms of released reducing sugar (g/L). Observed values were well fitted with second order polynomial equation (R2 = 0.91) and predicted outcomes were in agreement with experimental results. Optimized treatment conditions were: 36 °C, 31 days residence time and 81% moisture. A 2.9 fold increase in reducing sugar was released when substrates were treated under these conditions. Our research findings emphasize importance of optimizing biological treatment conditions to achieve high biogas yield when using wheat and pearl millet straw as feedstocks and similar treatment could be carried out for other lignocellulosic feedstocks.

The screening and identification of the biological control fungi Chaetomium spp. against wheat common root rot.

Bipolaris sorokiniana is a soil-borne fungal pathogen causing common root rot in wheat, and is difficult to control through chemical and agricultural means. Chaetomium spp. have been documented as potential biological control agents for plant diseases. In this study, seven Chaetomium strains were screened for their abilities to control B. sorokiniana on agar plates. Strain 22-10 significantly inhibited the growth of B. sorokiniana on potato dextrose agar plates, up to 66.7%. Its filtrate of liquid culture also inhibited the mycelial growth of B. sorokiniana, indicating that strain 22-10 produced secondary metabolites against B. sorokiniana. The incidence and disease indexes of common root rot significantly decreased in wheat after treatment with the crude extract of strain 22-10. Six active compounds were purified from crude extract of this fungus culture against B. sorokiniana. Chaetoviridin A showed the highest efficiency to inhibit the growth of B. sorokiniana. Strain 22-10 was identified as Chaetomium globosum based on phylogenetic analysis of the rDNA internal transcribed spacer region and microscopic characteristics. The high control efficiency of strain 22-10 of C. globosum against B. sorokiniana suggested it has potential to be a biocontrol agent for B. sorokiniana.

Asymmetry in the function and dynamics of the cytosolic group II chaperonin CCT/TRiC.

The eukaryotic group II chaperonin, the chaperonin-containing t-complex polypeptide 1 (CCT), plays an important role in cytosolic proteostasis. It has been estimated that as much as 10% of cytosolic proteins interact with CCT during their folding process. CCT is composed of 8 different paralogous subunits. Due to its complicated structure, molecular and biochemical investigations of CCT have been difficult. In this study, we constructed an expression system for CCT from a thermophilic fungus, Chaetomium thermophilum (CtCCT), by using E. coli as a host. As expected, we obtained recombinant CtCCT with a relatively high yield, and it exhibited fairly high thermal stability. We showed the advantages of the overproduction system by characterizing CtCCT variants containing ATPase-deficient subunits. For diffracted X-ray tracking experiment, we removed all surface exposed cysteine residues, and added cysteine residues at the tip of helical protrusions of selected two subunits. Gold nanocrystals were attached onto CtCCTs via gold-thiol bonds and applied for the analysis by diffracted X-ray tracking. Irrespective of the locations of cysteines, it was shown that ATP binding induces tilting motion followed by rotational motion in the CtCCT molecule, like the archaeal group II chaperonins. When gold nanocrystals were attached onto two subunits in the high ATPase activity hemisphere, the CtCCT complex exhibited a fairly rapid response to the motion. In contrast, the response of CtCCT, which had gold nanocrystals attached to the low-activity hemisphere, was slow. These results clearly support the possibility that ATP-dependent conformational change starts with the high-affinity hemisphere and progresses to the low-affinity hemisphere.

Architecture and mechanism of the late endosomal Rab7-like Ypt7 guanine nucleotide exchange factor complex Mon1-Ccz1.

The Mon1-Ccz1 complex (MC1) is the guanine nucleotide exchange factor (GEF) for the Rab GTPase Ypt7/Rab7 and is required for endosomal maturation and fusion at the vacuole/lysosome. Here we present the overall architecture of MC1 from Chaetomium thermophilum, and in combining biochemical studies and mutational analysis in yeast, we identify the domains required for catalytic activity, complex assembly and localization of MC1. The crystal structure of a catalytic MC1 core complex bound to Ypt7 provides mechanistic insight into its function. We pinpoint the determinants that allow for a discrimination of the Rab7-like Ypt7 over the Rab5-like Vps21, which are both located on the same membrane. MC1 shares structural similarities with the TRAPP complex, but employs a novel mechanism to promote nucleotide exchange that utilizes a conserved lysine residue of Ypt7, which is inserted upon MC1 binding into the nucleotide-binding pocket of Ypt7 and contributes to specificity.

The fungal endophyte Chaetomium globosum negatively affects both above- and belowground herbivores in cotton.

Mutualistic plant-endophyte symbioses can benefit plants by increasing host fitness through reductions in herbivory. The fungus, Chaetomium globosum strain TAMU 520, was previously isolated as an endophyte from cotton (Gossypium hirsutum) and can be re-inoculated to systemically colonize cotton plants via seed treatment. We evaluated the potential impacts of the endophyte in cotton on plant parasitic nematodes belowground, along with piercing-sucking and chewing insects aboveground. Endophytic C. globosum inhibited root-knot nematode (Meloidogyne incognita) infection and reduced female reproduction belowground. To confirm the endophytic effect of C. globosum on root-knot nematode, a contact fungicide was applied to remove soil-borne and epiphytic C. globosum Consistent inhibition of nematode activity was observed post-fungicide treatment, with positive C. globosum colonization confirmed within plant tissues. Aboveground, endophytic C. globosum also negatively affected the fecundity of both cotton aphids (Aphis gossypii) and beet armyworms (Spodoptera exigua). Faster development rates and smaller head capsule of beet armyworm larvae were observed when fed Chaetomium-colonized plants. However, no larval weight difference was found between Chaetomium-colonized and control plants. No consistent effect on plant performance was found across experiments. Our findings illustrate how a single facultative fungal endophyte can increase plant systemic resistance against a range of invertebrate herbivores in a major crop.

Infection by a foliar endophyte elicits novel arabidopside-based plant defence reactions in its host, Cirsium arvense.

Endophytic fungi live asymptomatically within plants. They are usually regarded as nonpathogenic or even mutualistic, but whether plants respond antagonistically to their presence remains unclear, particularly in the little-studied associations between endophytes and nongraminoid herbaceous plants. We investigated the effects of the endophyte Chaetomium cochlioides on leaf chemistry in Cirsium arvense. Plants were sprayed with spores; leaf material from both subsequent new growth and the sprayed leaves was analysed 2 wk later. Infection frequency was 91% and 63% for sprayed and new growth, respectively, indicating that C. cochlioides rapidly infects new foliage. Metabolomic analyses revealed marked changes in leaf chemistry with infection, especially in new growth. Changes in several novel oxylipin metabolites were detected, including arabidopsides reported here for the first time in a plant species other than Arabidopsis thaliana, and a jasmonate-containing galactolipid. The production of these metabolites in response to endophyte presence, particularly in newly infected foliage, suggests that endophytes elicit similar chemical responses in plants to those usually produced following wounding, herbivory and pathogen invasion. Whether endophytes benefit their hosts may depend on a complex series of chemically mediated interactions between the plant, the endophyte, other microbial colonists and natural enemies.

Quantification of C. globosum spores in house dust samples.

Chaetomium globosum is one of the most common fungi that grows in damp buildings and occurs in agricultural and forestry workplaces. Using sera from atopic patients, we characterized and purified an extracellular chitosanase (Chg47) from C. globosum that is antigenic to humans. The study reports the production of monoclonal antibodies to the protein. Three capture ELISAs were developed for Chg47 for detection of spores and spore and mycelial fragments in dust samples using different mono- and polyclonal antibody combinations. One method is based on an enhanced biotinylated polyclonal antibody as the secondary antibody and coating anti-IgM to capture one of two clones of IgM monoclonal antibodies as the capture antibody. The other method makes use of an enhanced rabbit polyclonal antibody as both the primary and capture antibody. The detection limit of the double PAb method for the Chg47 antigen was 7.6 pg/ml. When the anti-IgM+10B3 clone was used, the detection limit was 61 pg/ml and for anti-IgM+5F12, 122 pg/ml. The detection limit of double PAb method is comparable to methods for the allergen and spores of Aspergillus versicolor in house dust and is more sensitive than other immunoassays for allergens in house including for Stachybotrys chartarum, Aspergillus fumigatus and Alternaria alternata. All three methods had limited cross-reactivity to fungi common in house dust representing a diverse array of taxa.

Identification and characterization of rhizosphere fungal strain MF-91 antagonistic to rice blast and sheath blight pathogens.

AIM: To examine the inhibition effects of rhizosphere fungal strain MF-91 on the rice blast pathogen Magnaporthe grisea and sheath blight pathogen Rhizoctonia solani. METHODS AND RESULTS: Rhizosphere fungal strain MF-91 and its metabolites suppressed the in vitro mycelial growth of R. solani. The inhibitory effect of the metabolites was affected by incubation temperature, lighting time, initial pH and incubation time of rhizosphere fungal strain MF-91. The in vitro mycelial growth of M. grisea was insignificantly inhibited by rhizosphere fungal strain MF-91 and its metabolites. The metabolites of rhizosphere fungal strain MF-91 significantly inhibited the conidial germination and appressorium formation of M. grisea. Moreover, the metabolites reduced the disease index of rice sheath blight by 35·02% in a greenhouse and 57·81% in a field as well as reduced the disease index of rice blast by 66·07% in a field. Rhizosphere fungal strain MF-91 was identified as Chaetomium aureum based on the morphological observation, the analysis of 18S ribosomal DNA internal transcribed spacer sequence and its physiological characteristics, such as the optimal medium, temperature and initial pH for mycelial growth and sporulation production. CONCLUSIONS: Rhizosphere fungus C. aureum is effective in the biocontrolling of rice blast pathogen M. grisea and sheath blight pathogen R. solani both in in vitro and in vivo conditions. SIGNIFICANCE AND IMPACT OF THE STUDY: This study is the first to show that rhizosphere fungus C. aureum is a potential fungicide against rice blast and sheath blight pathogens.

Endophytic Chaetomium globosum enhances maize seedling copper stress tolerance.

This study aims at characterisation of the impact of Chaetomium globosum on copper stress resistance of maize seedlings. Higher levels of copper treatment decreased maize dry weight and induced a marked increase in osmotic solutes, antioxidant enzyme activity and the level of lipid peroxidation. On the other hand, addition of the endophytic C. globosum alleviated the toxic effect of copper on maize growth. The combination of copper sulphate and Chaetomium increased seedling dry weight, osmotic solute content and antioxidant enzyme activity compared to copper sulphate alone, while lipid peroxidation levels were also decreased. The fungal scavenger system might be important for supporting the ability of maize seedlings to resist copper toxicity.

Controlling Myzus persicae with recombinant endophytic fungi Chaetomium globosum expressing Pinellia ternata agglutinin: using recombinant endophytic fungi to control aphids.

AIMS: Sap-sucking insect pests have become the major threats to many crops in recent years; however, only a few biopesticides have been developed for controlling those pests. Here, we developed a novel pest management strategy, which uses endophytes to express anti-pest plant lectins. METHODS AND RESULTS: The fungal endophyte of Chaetomium globosum YY-11 with anti-fungal activities was isolated from rape seedlings. Pinellia ternata agglutinin (pta) gene was cloned into YY-11 mediated by Agrobacterium tumefaciens. The positive transformants, as selected by antibiotic resistance, were evaluated using PCR and Western blot assay. We found that the recombinant endophytes colonized most of the crops, and the resistance of rape inoculated with recombinant endophytic fungi significantly inhibited the growth and reproduction of Myzus persicae. CONCLUSIONS: Our results showed that the recombinant endophytes expressing Pinellia ernata agglutinin (PTA) may endow hosts with resistance against sap-sucking pests. SIGNIFICANCE AND IMPACT OF THE STUDY: This research may have important implications for using endophytes to deliver insecticidal plant lectin proteins to control sap-sucking pests for crop protection.

Cloning and characterization of a novel chitinase gene (chi46) from Chaetomium globosum and identification of its biological activity.

Chitinases play a major role in the defensive strategies of plants against fungal pathogens. In the current study, the gene for a 46-kDa endochitinase (chi46) was cloned from Chaetomium globosum, an important biocontrol fungus. The corresponding complementary deoxyribonucleic acid sequence was 1,350 bp in length, encoding 449 amino acid residues. The temporal expression of chi46, in response to the treatments of cell walls of six pathogens and confrontation against two fungal pathogens, was measured in C. globosum using real-time reverse transcription polymerase chain reaction. The expression of chi46 can be highly induced by exposure to the cell walls of plant pathogens and living pathogens, suggesting a role in plant disease resistance. The chi46 gene was inserted into the pPIC9 vector and transferred into the cells of Pichia pastoris GS115 for heterologous expression. The optimal reaction conditions for chitinase CHI46 activity were: 45 degrees C, pH of 5.0, and 5 mmol l(-1) of Cu2+. The maximum enzyme activity was 1.42 U ml(-1) following exposure to the cell wall chitin of Septoria tritici. The CHI46 enzyme can efficiently degrade cell walls of the phytopathogenic Rhizoctonia solani, Fusarium oxysporum, Sclerotinia sclerotiorum, Valsa sordida, S. tritici, and Phytophthora sojae, demonstrating that it may be involved in the biocontrol mechanism of C. globosum.

[Adaptation of Chaetomium globosum 3250 Kunze ex Fr. to the rhizosphere of spring wheat and its ability to colonize the root system].

The ability of the natural strain of marsupial fungus Chaetomium globosum 3250 to adapt to the rhizosphere soil of the spring wheat, and to colonize its root system has been studied in the laboratory and field experiments. It is established that C. globosum 3250 adapts to the rhyzosphere soil of culture during vegetative period, actively colonizes the culture root system, forcing the agents of roots rots out of it. A preparation of Khetomyk is created on the basis of the above culture for the improvement of plants feeding and their defence from the agents of root diseases. When applying Khetomyk on sowings of the spring wheat a considerable limitation of development of root rots took place (the disease distribution decreased 2.9 times, and intensity of display--3.9 times), as well as the improvement of plants' feeding, increase of chlorophyll a and b maintenance in leafs. All these provided the considerable increase of the spring wheat harvest--by 30.1% as compared to the control. The culture harvest structure analysis revealed positive influence of Chetomyk on the ear length, the number of seeds in the ear, mass of the seeds in the ear. In experiments with the spring wheat grown in sterile conditions, the ability of C. globosum 3250 to penetrate inside in rhyzoderma cells and root fuzzes and to form carposomes on the root surface has been shown.

A heat shock protein gene (hsp22.4) from Chaetomium globosum confers heat and Na2CO3 tolerance to yeast.

A small heat shock protein gene (hsp22.4) was cloned from Chaetomium globosum using rapid amplification of cDNA ends (RACE). The 986-bp full-length hsp22.4 cDNA contains a 609-bp open reading frame encoding a 202-amino-acid protein with an estimated molecular mass of 22.4 kDa. The hsp22.4 gene was amplified using specific primers in the 5' and 3' untranslated regions of the hsp22.4 cDNA. The temporal expression of hsp22.4 was measured in C. globosum by real-time reverse transcriptase-polymerase chain reaction after exposure to heat, cold, Na(2)CO(3), and NaCl. The expression of hsp22.4 was induced by heat and Na(2)CO(3) treatment and inhibited by cold and NaCl treatment. The hsp22.4 gene was inserted into pYES2 containing the inducible GAL1 promoter and transferred into yeast (Saccharomyces cerevisiae) for expression. The hsp22.4 transgenic yeast displayed significantly greater resistance to heat and Na(2)CO(3) stresses than control (yeast cells transformed with empty pYES2), suggesting that the expression of hsp22.4 gene confers not only heat tolerance but also significant alkali (Na(2)CO(3)) stress tolerance.

Cellobiose dehydrogenase formation by filamentous fungus Chaetomium sp. INBI 2-26(-).

Laccase-negative filamentous fungus INBI 2-26(-) isolated from non-sporulating laccase-forming fungal association INBI 2-26 by means of protoplast technique was identified as Chaetomium sp. based on partial sequence of its rRNA genes. In the presence of natural cellulose sources, the strain secreted neutral cellobiose dehydrogenase (CDH) activity both in pure culture and in co-culture with laccase-positive filamentous fungus INBI 2-26(+) isolated from the same association. INBI 2-26(-) also secreted CDH during submerged cultivation in minimal medium with glucose as the sole carbon source. Maximal CDH activity of 1IU/ml at pH 6 with 2,6-dichlorophenolindophenol (DCPIP) as an acceptor was obtained on 12th day of submerged cultivation with filter paper as major cellulose source. Cellulase system of Chaetomium sp. INBI 2-26(-) capable of adsorption onto H(3)PO(4)-swollen filter paper consisted of four major proteins (Mr 200, 95, 65 and 55K) based on SDS-polyacrylamide gel electrophoresis and was capable of DCPIP reduction without exogenous cellobiose.